Proteome Profiler Human XL Oncology Array Best Seller
R&D Systems, part of Bio-Techne | Catalog # ARY026
Key Product Details
Summary for Proteome Profiler Human XL Oncology Array
To detect differences in 84 cancer-related proteins between samples. No specialized equipment is necessary.
- Troubleshooting Guide
- Assays for Analytes represented in the Human XL Oncology Array Kit
- Compatible with LI-COR* and chemiluminescence detection
- LI-COR Data Examples for the Human Oncology Array Kit
General Assay Principle
Carefully selected capture antibodies have been spotted in duplicate on nitrocellulose membranes. Cell culture supernates, cell lysates, tissue lysates, serum, plasma, urine, saliva, or human milk are diluted and incubated with the Human XL Oncology Array overnight. The array is washed to remove unbound proteins, followed by incubation with a cocktail of biotinylated detection antibodies. Streptavidin-HRP and chemiluminescent detection reagents are applied, and a signal is produced at each capture spot corresponding to the amount of protein bound. Chemiluminescence is detected in the same manner as a Western blot.
- 4 Array Membranes
- 4-Well Multi-dish
- Array Buffers
- Wash Buffer
- Detection Antibody Cocktail
- Streptavidin-HRP
- Chemiluminescent Detection Reagents
- Transparency Overlay Template
- Detailed Protocol
For a complete list of the kit contents and necessary materials, please see the Materials Provided/Other Supplies Required sections of the product datasheet.
Stability and Storage
Reagents are stable for 12 months from date of receipt when stored in the dark at 2° C to 8°.
| Simultaneously detect the levels of these cancer-related proteins in a single sample. | ||
| AFP | ErbB4 | MMP-2 |
| Amphiregulin | FGF basic | MMP-3 |
| Angiopoietin-1 | FoxC2 | MMP-9 |
| ANGPTL4 | FKHR | MSP/MST1 |
| ENPP-2/Autotaxin | Galectin-3 | MUC-1 |
| AXL | GM-CSF | Nectin-4 |
| BCL-X | HCG | Osteopontin |
| CA125/MUC-16 | HGF R/c-Met | p27/Kip1 |
| E-Cadherin | HIF-1alpha | p53 |
| VE-Cadherin | HNF-3beta | PDGF-AA |
| CAP-G | HO-1/HMOX1 | CD31/PECAM-1 |
| CA-9 | ICAM-1/CD54 | Progesterone R |
| Cathepsin B | CD25/IL-2 R alpha | Progranulin |
| Cathepsin D | IL-6 | Prolactin |
| Cathepsin S | CXCL8/IL-8 | Prostasin |
| CEACAM-5 | IL-18 Bpa | E-Selectin |
| Decorin | KLK-3/PSA | Maspin |
| DKK-1 | KLK-5 | PAI-1/Serpin E1 |
| DLL-1 | KLK-6 | SNAIL |
| EGF R/ErbB1 | Leptin (OB) | SPARC |
| Endoglin/CD105 | Lumican | Survivin |
| Endostatin | CCL2/MCP-1 | Tenascin-C |
| Enolase 2 | CCL8/MCP-2 | THBS-1 |
| eNOS | CCL7/MCP-3 | TIE-2 |
| EpCAM | M-CSF | UPA-1 |
| ER-alpha | Mesothelin | VCAM-1 |
| ErbB2 | CCL3/MIP-1alpha | VEGF |
| ErbB3 | CCL20/MIP-3alpha | Vimentin |
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, relative expression levels of human cancer-related proteins in samples can be determined using the following procedure:
- Prepare membrane and incubate with prepared sample
- Incubate the membrane with Detection Antibody Cocktail
- Incubate the membrane array with Streptavidin-HRP
- Develop the membrane array with Chemi Reagents 1 and 2
- Expose the membrane array to autoradiography film
- Rectangular 4-Well Multi-dish
- 4 Human XL Oncology Array nitrocellulose membranes spotted with 84 different antibodies to cancer-related proteins
- Array Buffer 4
- Array Buffer 6
- Chemi Reagent 1
- Chemi Reagent 2
- Detection Antibody Cocktail, Human XL Oncology Array
- Streptavidin-HRP
- Transparency Overlay Template
- Wash Buffer Concentrate (25X)
Reagents
- Pipettes and pipette tips
- Gloves
- Plastic container with the capacity to hold 50 mL (for washing the arrays)
- Plastic transparent sheet protector (trimmed to 10 cm x 12 cm and open on three sides)
- Plastic wrap
- Absorbent lab wipes (KimWipes® or equivalent)
- Paper towels
- X-ray film (Kodak BioMax™ Light-1) or equivalent
- Flat-tipped tweezers
Equipment
- Rocking platform shaker
- Microcentrifuge
- Autoradiography cassette
- Film developer
- Flatbed scanner with transparency adapter capable of transmission mode
- Computer capable of running image analysis software and Microsoft Excel
Other Supplies Required for Cell Lysate Samples
- Phosphate-Buffered Saline (PBS)
- Lysis buffer 17
- Aprotinin
- Leupeptin (Catalog # EI002)
- Pepstatin (Catalog # EI003)
Other Supplies Required for Tissue Lysate Samples
- Protease Inhibitor Cocktail
- Igepal® CA-630
- Sodium deoxycholate
- Sodium dodecyl sulfate
R&D Systems Protocol for Multiple Analyte Detection Using the Proteome Profiler™ Human XL Oncology Array Kit, Panel A (Catalog # ARY026)
Add 2 mL of Array Buffer 6 to each well of the supplied 4-Well Multi-dish.
Place each array membrane in a separate well of the 4-Well Multi-dish
Incubate for one hour on a rocking platform shaker.
Add 0.5 mL Array Buffer 4 to each sample.
Adjust volume of each sample to final volume of 1.5 mL with Array Buffer 6.
Replace the Array Buffer 6 in each well of the 4-Well Multi-dish with prepared samples.
Incubate overnight at 2 °C to 8 °C on a rocking platform.
Wash each array membrane 3 times with 1X Wash Buffer in a separate container.
Wash each well of the 4-Well Multi-dish with 1X Wash Buffer.
Add 30 µL of Detection Antibody Cocktail to 1.5 mL of Array Buffer 4/6 for each array.
Pipette 1.5 mL diluted Detection Antibody Cocktail into each well of the 4-Well Multi-dish.
Return each array membrane to the 4-Well Multi-dish containing diluted Detection Antibody Cocktail.
Incubate for one hour on a rocking platform shaker.
Wash each array membrane 3 times with 1X Wash Buffer in a separate container.
Wash each well of the 4-Well Multi-dish with 1X Wash Buffer.
Add 2 mL of diluted Streptavidin-HRP to each well of the 4-Well Multi-dish.
Place the array membrane in the diluted Streptavidin-HRP solution.
Incubate for 30 minutes on a rocking platform shaker.
Wash each array membrane 3 times with 1X Wash Buffer in a separate container.
Place the array membrane on a plastic sheet protector.
Pipette 1 mL of the prepared Chemi Reagent Mix evenly onto the membrane.
Cover the membrane with the top sheet of the plastic protector
Incubate for 1 minute.
Blot off excess Chemi Reagent Mix.
Wrap the membrane and sheet protector in plastic wrap.
Place the wrapped array membrane in an autoradiography film cassette and expose to X-ray film.
